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1.
J Fish Dis ; 45(1): 59-68, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34536027

RESUMO

White spot syndrome virus (WSSV) is a pathogenic and threatening virus in shrimp culture for which there is no effective control strategy. Finding antiviral lead compounds for the development of anti-WSSV drugs is urgent and necessary; in this study, esculin from 12 monomeric compounds exhibited an excellent anti-WSSV activity. The results showed that esculin increased the survival rate of WSSV-infected shrimps by 59% and reduced the virus copy number in vivo over 90% at 100 µM. In the pre-treatment and post-treatment experiments, esculin could prevent and treat WSSV infection. Compared with the control group, the virus copy number decreased by 30% after 6 h of esculin pre-incubation with WSSV particles and inhibited horizontal transmission of WSSV to a certain extent. Considering that the antiviral activity of esculin was stable in the aquacultural water for 2 days, we evaluated the dosing pattern of continuous medication changes. Obviously, the survival rate of WSSV-infected shrimps was 0% at 108 h when no esculin exchange was made, while at 120 h the survival rate was over 40% at continuous medicine changes. In addition, esculin significantly increased the expression of antimicrobial peptides and thus improved the ability of shrimp to resist WSSV. Overall, our findings suggest that esculin has the potential to be developed into an anti-WSSV medicine.


Assuntos
Antivirais/farmacologia , Esculina/farmacologia , Doenças dos Peixes , Penaeidae , Vírus da Síndrome da Mancha Branca 1 , Animais , Aquicultura , Surtos de Doenças , Doenças dos Peixes/tratamento farmacológico , Doenças dos Peixes/virologia , Penaeidae/virologia , Vírus da Síndrome da Mancha Branca 1/efeitos dos fármacos
2.
Phytomedicine ; 92: 153687, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34482222

RESUMO

BACKGROUND: Aesculin (AES), an effective component of Cortex fraxini, is a hydroxycoumarin glucoside that has diverse biological properties. The nucleotide-binding domain leucine-rich repeat-containing receptor, pyrin domain-containing 3 (NLRP3) inflammasome has been heavily interwoven with the development of myocardial ischemia/reperfusion injury (MIRI). Nevertheless, it remains unclear whether AES makes a difference to the changes of the NLRP3 inflammasome in MIRI. PURPOSE: We used rats that were subjected to MIRI and neonatal rat cardiomyocytes (NRCMs) that underwent oxygen-glucose deprivation/restoration (OGD/R) process to investigate what impacts AES exerts on MIRI and the NLRP3 inflammasome activation. METHODS: The establishment of MIRI model in rats was conducted using the left anterior descending coronary artery ligation for 0.5 h ischemia and then untying the knot for 4 h of reperfusion. After reperfusion, AES were administered intraperitoneally using 10 and 30 mg/kg doses. We evaluated the development of reperfusion ventricular arrhythmias, hemodynamic changes, infarct size, and the biomarkers in myocardial injury. The inflammatory mediators and pyroptosis were also assessed. AES at the concentrations of 1, 3, and 10 µM were imposed on the NRCMs immediately before the restoration process. We also determined the cell viability and cell death in the NRCMs exposed to OGD/R insult. Furthermore, we also analyzed the levels of proteins that affect the NLRP3 inflammasome activation, pyroptosis, and the AKT serine/threonine kinase (Akt)/glycogen synthase kinase 3 beta (GSK3ß)/nuclear factor-kappa B (NF-κB) signaling pathway via western blotting. RESULTS: We found that AES notably attenuated reperfusion arrhythmias and myocardia damage, improved the hemodynamic function, and ameliorated the inflammatory response and pyroptosis of cardiomyocytes in rats and NRCMs. Additionally, AES reduced the NLRP3 inflammasome activation in rats and NRCMs. AES also enhanced the phosphorylation of Akt and GSK3ß, while suppressing the phosphorylation of NF-κB. Moreover, the allosteric Akt inhibitor, MK-2206, abolished the AES-mediated cardioprotection and the NLRP3 inflammasome suppression. CONCLUSIONS: These findings indicate that AES effectively protected cardiomyocytes against MIRI by suppressing the NLRP3 inflammasome-mediated pyroptosis, which may relate to the upregulated Akt activation and disruption of the GSK3ß/NF-κB pathway.


Assuntos
Inflamassomos , Traumatismo por Reperfusão Miocárdica , Animais , Esculina , Quinase 3 da Glicogênio Sintase , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Proteína 3 que Contém Domínio de Pirina da Família NLR , Proteínas Proto-Oncogênicas c-akt , Piroptose , Ratos
3.
J Biotechnol ; 337: 46-56, 2021 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-34197823

RESUMO

Esculin is a polyphenol with multiple bioactivities and poor lipophilicity. Therefore, a whole-cell catalytic strategy for esculin acylation was developed to improve its lipophilicity. A total of 12 strains were tested, among which Pseudomonas stutzeri exhibited the highest catalytic activity and mono-acylated regioselectivity. The conversion reached the highest level of 92.7 % at 24 h under the optimal conditions, when vinyl acetate was used as an acyl donor. The catalytic ability of P. stutzeri remained above 60 % after three cycles. Subsequently, five esculin esters with different lengths of fatty chains were synthesized and structurally identified. Of them, esculin-6'-O-octanoate, esculin-6'-O-laurate, and esculin-6'-O-myristate exhibited cytotoxicity on LO2 cells by inducing apoptosis and necrosis. The cytotoxicity of these three esters may attribute to their membrane-disrupting properties. This study provides a novel whole-cell biocatalytic strategy for the acylation of esculin and insight for application of esculin esters as a food additive or drug.


Assuntos
Esculina , Ésteres , Acilação , Biocatálise , Lipase/metabolismo
4.
Mater Sci Eng C Mater Biol Appl ; 118: 111397, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33255002

RESUMO

Moderate and prolonged payload release in response to a particular factor is highly demanded for efficient carriers of low-molecular-weight, chemically unstable phytopharmaceuticals. Thus, the objective of our contribution was to establish the effect of pH-responsive polyelectrolyte coatings on the release properties of carboxymethyl cellulose-based microparticles designed to deliver phytopharmaceuticals through the gastrointestinal tract. Microparticles were fabricated via extrusion coupled with external gelation and further coated with polyelectrolytes (PEs) (chitosan, gelatin, or PAH and PSS) involving electrostatic interactions. Successful deposition of PEs was confirmed by FTIR, and their thickness and viscosity were characterized in terms of QCM-D and ellipsometric techniques. The encapsulation efficiency of esculin, used as a model phytopharmaceutical, as proven by UV-Vis studies, was over 57%. SEM and fluorescence microscopy revealed a micrometric size, a mostly spherical shape and an altered topography of the investigated microcapsules. The physical stability of the microcapsules in media of various pH values was confirmed with CLSM and gravimetric studies. Studies on human gingival fibroblasts in vitro revealed that the obtained microparticles did not induce any cytotoxic effects. Payload release was monitored in situ by means of CLSM and ex situ under gastrointestinal conditions in vitro. Mathematical evaluation of the microparticle release profiles using classical models led to the establishment of a new hybrid model that revealed the mechanism behind esculin release. We demonstrated that the application of a polyelectrolyte shell onto CMC-based microspheres may provide controlled delivery of the payload, with its release triggered by the pH and ionic strength of the medium. These observations suggest that the release manner of small-molecule glycosides under gastrointestinal conditions can be tailored by careful selection of suitable materials to obtain biocompatible and functional hydrogel microparticles.


Assuntos
Carboximetilcelulose Sódica , Quitosana , Preparações de Ação Retardada , Esculina , Humanos , Concentração de Íons de Hidrogênio , Microesferas , Polieletrólitos
5.
Molecules ; 25(21)2020 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-33171577

RESUMO

Coumarins, which occur naturally in the plant kingdom, are diverse class of secondary metabolites. With their antiproliferative, chemopreventive and antiangiogenetic properties, they can be used in the treatment of cancer. Their therapeutic potential depends on the type and location of the attachment of substituents to the ring. Therefore, the aim of our study was to investigate the effect of simple coumarins (osthole, umbelliferone, esculin, and 4-hydroxycoumarin) combined with sorafenib (specific inhibitor of Raf (Rapidly Accelerated Fibrosarcoma) kinase) in programmed death induction in human glioblastoma multiforme (T98G) and anaplastic astrocytoma (MOGGCCM) cells lines. Osthole and umbelliferone were isolated from fruits: Mutellina purpurea L. and Heracleum leskowii L., respectively, while esculin and 4-hydroxycoumarin were purchased from Sigma Aldrich (St. Louis, MO, USA). Apoptosis, autophagy and necrosis were identified microscopically after straining with specific fluorochromes. The level of caspase 3, Beclin 1, PI3K (Phosphoinositide 3-kinase), and Raf kinases were estimated by immunoblotting. Transfection with specific siRNA (small interfering RNA) was used to block Bcl-2 (B-cell lymphoma 2), Raf, and PI3K expression. Cell migration was tested with the wound healing assay. The present study has shown that all the coumarins eliminated the MOGGCCM and T98G tumor cells mainly via apoptosis and, to a lesser extent, via autophagy. Osthole, which has an isoprenyl moiety, was shown to be the most effective compound. Sorafenib did not change the proapoptotic activity of this coumarin; however, it reduced the level of autophagy. At the molecular level, the induction of apoptosis was associated with a decrease in the expression of PI3K and Raf kinases, whereas an increase in the level of Beclin 1 was observed in the case of autophagy. Inhibition of the expression of this protein by specific siRNA eliminated autophagy. Moreover, the blocking of the expression of Bcl-2 and PI3K significantly increased the level of apoptosis. Osthole and sorafenib successfully inhibited the migration of the MOGGCCM and T98G cells.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Cumarínicos/farmacologia , Glioblastoma/tratamento farmacológico , Magnoliopsida/química , Extratos Vegetais/farmacologia , Sorafenibe/farmacologia , 4-Hidroxicumarinas/farmacologia , Antineoplásicos Fitogênicos/isolamento & purificação , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Proteína Beclina-1/genética , Proteína Beclina-1/metabolismo , Caspase 3/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Esculina/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Necrose/tratamento farmacológico , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Interferente Pequeno/metabolismo , Umbeliferonas/farmacologia , Quinases raf/metabolismo
6.
J Affect Disord ; 277: 755-764, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33065814

RESUMO

BACKGROUND: The purpose of this study was to evaluate whether Esculin could improve the depressive symptom induced by LPS in mice and explore the role of CCR5 in its potential mechanism. METHODS: Mice were stimulated with LPS to establish depression model and treated with Esculin. The emotional alteration was assessed via behavior tests. The ELISA assay and western blot analysis were applied to detect the expressions of inflammatory cytokines and correlative proteins. RESULTS: As a result, Esculin played a protective role in LPS-induced depressive dysfunction, which was possible through the reduction of M1 microglia, and elevation of M2 microglia by inhibiting TLR4/NF-κB signaling pathway regulated by CCR5. Besides, Esculin led to up-regulation of the CREB/BDNF neuroprotective pathway, and suppression of inflammatory cytokines both in the central and peripheral system. BV2 cells were stimulated with LPS to further elucidate the accordant mechanism in vitro. Molecular docking results suggested that Esc bound to CCR5 at amino acid residues TYR187 and THR105 through hydrogen-bonding. LIMITATIONS: Transgenic animals might be useful for the further investigation. CONCLUSIONS: From the overall results, we concluded that Esculin might exert a beneficial effect on LPS-induced depression in mice and represent an effective treatment for depression.


Assuntos
Esculina , Lipopolissacarídeos , Animais , Citocinas , Depressão/induzido quimicamente , Depressão/tratamento farmacológico , Lipopolissacarídeos/toxicidade , Camundongos , Simulação de Acoplamento Molecular , NF-kappa B
7.
Food Funct ; 11(10): 9129-9143, 2020 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-33026011

RESUMO

Herein, a sturgeon skin gelatine film combined with esculin and ferric citrate was developed as an edible food packaging material to prevent Enterococcus faecalis (E. faecalis) contamination. E. faecalis is able to hydrolyse esculin in the film, and then the hydrolysed product, esculetin, combines with ferric citrate to form a brown-black phenol iron complex. This phenomenon can be observed easily after 48 h of contamination under visible light, and it can be determined under 365 nm ultraviolet light with high sensitivity. With the addition of esculin and ferric citrate, the film showed better mechanical properties and water vapour permeability than those of the unmodified gelatine. When an increased amount of esculin was added, an increase in thermal stability, antioxidant activity, and antioxidant stability of the film was observed. These physicochemical characteristics are beneficial for developing a packaging material for food storage that mitigates foodborne illness caused by E. faecalis.


Assuntos
Esculina/química , Compostos Férricos/química , Proteínas de Peixes/química , Embalagem de Alimentos/instrumentação , Gelatina/química , Pele/química , Animais , Antibacterianos/química , Antibacterianos/farmacologia , Antioxidantes/química , Antioxidantes/farmacologia , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/fisiologia , Esculina/farmacologia , Peixes , Contaminação de Alimentos/prevenção & controle , Doenças Transmitidas por Alimentos/microbiologia , Doenças Transmitidas por Alimentos/prevenção & controle , Humanos
8.
Zhonghua Zhong Liu Za Zhi ; 42(8): 629-634, 2020 Aug 23.
Artigo em Chinês | MEDLINE | ID: mdl-32867453

RESUMO

Objective: To investigate the effect of esculin on the proliferation of triple negative breast cancer cells and its molecular mechanism. Methods: MDA-MB-231 cells were treated with 28, 56, 112, 225, 450 and 900 µmol/L of esculin for 24, 48 and 72 h, respectively, and the cell viability was detected by cell counting kit 8 (CCK-8) assay. In addition, MDA-MB-231 cells were treated with 0, 225, 450 and 900 µmol/L of esculin for 48 h. And then the changes in cell morphology were observed by inverted microscope. The clone-forming ability was detected by colony formation assay. The mRNA expression levels of FBI-1, p53 and p21 were detected using real-time fluorescence quantitative polymerase chain reaction. The protein expression levels of FBI-1, p53, p21 and Ki67 were detected by western blot. Results: Compared with the blank control group, the cell viability of MDA-MB-231 cells that treated with esculin significantly decreased in a dose-dependent and time-dependent manners. After treatment with esculin, MDA-MB-231 cells shrunk, flattened, adhered poorly to the culture dish and the cell spacing became larger. Meanwhile, shedding and incomplete cells appeared, of which 900 µmol/L of esculin treatment group showed the most dramatic changes. In addition, the colony formation ratios were decreased to (77.18±5.13)%, (65.94±4.98)% and (45.92±3.70)% in the 225, 450 and 900 µmol/L of esculin treatment groups compared with blank control, respectively (P<0.01). Furthermore, the mRNA and protein expressions of FBI-1 increased, while the levels of p53 and p21 mRNA and protein, as well as the protein expression of Ki67 decreased in a concentration-dependent manner (P<0.01). Conclusion: Esculin may regulate cell cycle-related p53-p21 pathway via FBI-1 mediated DNA replication, thus inhibit the proliferation of triple negative breast cancer cells.


Assuntos
Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Esculina/farmacologia , RNA Mensageiro/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Neoplasias da Mama/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Proteínas de Ligação a DNA , Regulação para Baixo/efeitos dos fármacos , Feminino , Humanos , Fatores de Transcrição , Neoplasias de Mama Triplo Negativas/patologia
9.
Plant Cell ; 32(10): 3206-3223, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32769131

RESUMO

During their first year of growth, overwintering biennial plants transport Suc through the phloem from photosynthetic source tissues to storage tissues. In their second year, they mobilize carbon from these storage tissues to fuel new growth and reproduction. However, both the mechanisms driving this shift and the link to reproductive growth remain unclear. During vegetative growth, biennial sugar beet (Beta vulgaris) maintains a steep Suc concentration gradient between the shoot (source) and the taproot (sink). To shift from vegetative to generative growth, they require a chilling phase known as vernalization. We studied sugar beet sink-source dynamics upon vernalization and showed that before flowering, the taproot underwent a reversal from a sink to a source of carbohydrates. This transition was induced by transcriptomic and functional reprogramming of sugar beet tissue, resulting in a reversal of flux direction in the phloem. In this transition, the vacuolar Suc importers and exporters TONOPLAST SUGAR TRANSPORTER2;1 and SUCROSE TRANSPORTER4 were oppositely regulated, leading to the mobilization of sugars from taproot storage vacuoles. Concomitant changes in the expression of floral regulator genes suggest that these processes are a prerequisite for bolting. Our data will help both to dissect the metabolic and developmental triggers for bolting and to identify potential targets for genome editing and breeding.


Assuntos
Beta vulgaris/fisiologia , Floema/metabolismo , Proteínas de Plantas/metabolismo , Brotos de Planta/metabolismo , Metabolismo dos Carboidratos , Dióxido de Carbono/metabolismo , Temperatura Baixa , Esculina/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Floema/genética , Fotossíntese/fisiologia , Proteínas de Plantas/genética , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Brotos de Planta/genética , Sacarose/metabolismo , Açúcares/metabolismo , Vacúolos/genética , Vacúolos/metabolismo
10.
Plant Cell ; 32(11): 3485-3499, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32843436

RESUMO

Developmental transitions in plants require adequate carbon resources, and organ abscission often occurs due to competition for carbohydrates/assimilates. Physiological studies have indicated that organ abscission may be activated by Suc deprivation; however, an underlying regulatory mechanism that links Suc transport to organ shedding has yet to be identified. Here, we report that transport of Suc and the phytohormone auxin to petals through the phloem of the abscission zone (AZ) decreases during petal abscission in rose (Rosa hybrida), and that auxin regulates Suc transport into the petals. Expression of the Suc transporter RhSUC2 decreased in the AZ during rose petal abscission. Similarly, silencing of RhSUC2 reduced the Suc content in the petals and promotes petal abscission. We established that the auxin signaling protein RhARF7 binds to the promoter of RhSUC2, and that silencing of RhARF7 reduces petal Suc contents and promotes petal abscission. Overexpression of RhSUC2 in the petal AZ restored accelerated petal abscission caused by RhARF7 silencing. Moreover, treatment of rose petals with auxin and Suc delayed ethylene-induced abscission, whereas silencing of RhARF7 and RhSUC2 accelerated ethylene-induced petal abscission. Our results demonstrate that auxin modulates Suc transport during petal abscission, and that this process is regulated by a RhARF7-RhSUC2 module in the AZ.


Assuntos
Flores/fisiologia , Ácidos Indolacéticos/metabolismo , Rosa/fisiologia , Sacarose/metabolismo , Transporte Biológico , Esculina/metabolismo , Etilenos/metabolismo , Etilenos/farmacologia , Regulação da Expressão Gênica de Plantas , Proteínas de Fluorescência Verde/genética , Ácidos Indolacéticos/farmacologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Rosa/efeitos dos fármacos , Sacarose/farmacologia
11.
Life Sci ; 254: 117787, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32417372

RESUMO

AIMS: To evaluate the effects of esculin treatment on P2X7 receptor and mitochondrial dysfunction in the renal cortex of diabetic rats. MAIN METHODS: Male Wistar rats, 7 weeks old, were unilaterally nephrectomized. Part of these animals were induced to diabetes using streptozotocin (60 mg/kg). Diabetes was confirmed 48 h after induction, with blood glucose levels ≥200 mg/dL. Part of control and diabetic animals were selected to receive daily doses of esculin (50 mg/kg), during 8 weeks. The animals were placed in metabolic cages at the eighth week of protocol for 24 h urine collection and a small aliquot of blood was collected for biochemical analysis. After this procedure, the animals were euthanized and the remaining kidney was stored for histopathological analysis, Western blotting and mitochondrial high-resolution respirometry. KEY FINDINGS: Although esculin did not change metabolic parameters, renal biochemical function, neither TBARS in DM rats, esculin reduced P2X7 levels in these animals and restored mitochondrial function via glycolysis substrates and ß-oxidation. Besides, at the histological analysis, we observed that esculin reduced inflammatory infiltrates and collagen IV deposits as compared to diabetic group. SIGNIFICANCE: Esculin attenuated the development of renal injuries caused by hyperglycemia, proinflammatory and oxidative mechanisms mediated by P2X7 receptor, as seen by histological findings and improved mitochondrial function in diabetic animals. This suggests that esculin could be used as an adjuvant therapy to prevent the diabetic nephropathy.


Assuntos
Diabetes Mellitus Experimental/metabolismo , Esculina/farmacologia , Córtex Renal/metabolismo , Mitocôndrias/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Animais , Glicemia/metabolismo , Diabetes Mellitus Experimental/patologia , Colágenos Fibrilares/metabolismo , Glicólise/efeitos dos fármacos , Inflamação/prevenção & controle , Córtex Renal/patologia , Masculino , Oxirredução/efeitos dos fármacos , Ratos , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo
12.
Sci Rep ; 10(1): 4413, 2020 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-32157138

RESUMO

Tuberculosis (TB) is a leading cause of death worldwide and its impact has intensified due to the emergence of multi drug-resistant (MDR) and extensively drug-resistant (XDR) TB strains. Protein phosphorylation plays a vital role in the virulence of Mycobacterium tuberculosis (M.tb) mediated by protein kinases. Protein tyrosine phosphatase A (MptpA) undergoes phosphorylation by a unique tyrosine-specific kinase, protein tyrosine kinase A (PtkA), identified in the M.tb genome. PtkA phosphorylates PtpA on the tyrosine residues at positions 128 and 129, thereby increasing PtpA activity and promoting pathogenicity of MptpA. In the present study, we performed an extensive investigation of the conformational behavior of the intrinsically disordered domain (IDD) of PtkA using replica exchange molecular dynamics simulations. Long-term molecular dynamics (MD) simulations were performed to elucidate the role of IDD on the catalytic activity of kinase core domain (KCD) of PtkA. This was followed by identification of the probable inhibitors of PtkA using drug repurposing to block the PtpA-PtkA interaction. The inhibitory role of IDD on KCD has already been established; however, various analyses conducted in the present study showed that IDDPtkA had a greater inhibitory effect on the catalytic activity of KCDPtkA in the presence of the drugs esculin and inosine pranobex. The binding of drugs to PtkA resulted in formation of stable complexes, indicating that these two drugs are potentially useful as inhibitors of M.tb.


Assuntos
Proteínas de Bactérias/metabolismo , Esculina/farmacologia , Inosina Pranobex/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Proteínas Tirosina Fosfatases/metabolismo , Proteínas Tirosina Quinases/metabolismo , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/química , Reposicionamento de Medicamentos , Esculina/química , Inosina Pranobex/química , Modelos Moleculares , Simulação de Dinâmica Molecular , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/patogenicidade , Fosforilação , Ligação Proteica/efeitos dos fármacos , Conformação Proteica , Domínios Proteicos , Proteínas Tirosina Fosfatases/química , Desdobramento de Proteína , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/química
13.
Food Chem ; 310: 125858, 2020 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-31753682

RESUMO

This work established a binary ionic liquid-solvent system for effective enzymatic esterification of naturally occurring phenolic glycosides (flavonoids); which could result in a dramatic enhancement of Novozym 435-catalyzed esterification of esculin, demonstrating a great synergetic effect. In essence, [OMIM][BF4]-toluene and [TOMA][Tf2N]-hexane binary systems both served >90 mol% of conversions of esculin after 96 h of reaction at 60 °C. Typically, binary [TOMA][Tf2N]-hexane system enabled Novozym 435 with extremely high catalytic efficiency (kcat/Km = 17.57 × 10-2 (Ms)-1), which was 55-fold higher than that Novozym 435 exhibited in t-butanol solvent (one of the best solvent systems for esterification reactions). It was also found that the superiormatching in property and structure between IL and solvent was the decisive factor for the outperformance of [TOMA][Tf2N]-hexane binary system, in which [TOMA] and hexane facilitate the solubilization of esculin and fatty acids and [Tf2N]- anions and hexane offer protective effects for lipase at elevated temperatures.


Assuntos
Esculina/química , Líquidos Iônicos/química , Solventes/química , Catálise , Enzimas Imobilizadas , Esterificação , Proteínas Fúngicas , Lipase , Viscosidade
14.
Food Res Int ; 123: 414-424, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31284993

RESUMO

Bunium species have been reported to be used both as food and in traditional medicines. The scientific community has attempted to probe into the pharmacological and chemical profiles of this genus. Nonetheless, many species have not been investigated fully to date. In this study, we determined the phenolic components, antimicrobial, antioxidant, and enzyme inhibitory activities of aerial parts of four Bunium species (B. sayai, B. pinnatifolium, B. brachyactis and B. macrocarpum). Results showed that B. microcarpum and B. pinnatifolium were strong antioxidants as evidenced in the DPPH, ABTS, CUPRAC, and FRAP assays. B. brachyactis was the most effective metal chelator, and displayed high enzyme inhibition against cholinesterase, tyrosinase, amylase, glucosidase, and lipase. The four species showed varied antimicrobial activity against each microorganism. Overall, they showed high activity against P. mirabilis and E. coli (MIC and MBC <1 mg mL-1). B. brachyactis was more effective against Aspergillus versicolor compared to the standard drug ketoconazole. B. brachyactis was also more effective than both ketoconazole and bifonazole against Trichoderma viride. B. sayai was more effective than ketoconazole in inhibiting A. fumigatus. B. sayai was most non-toxic to HEK 293 (cellular viability = 117%) and HepG2 (cellular viability = 104%). The highest level of TPC was observed in B. pinnatifolium (35.94 mg GAE g-1) while B. microcarpum possessed the highest TFC (39.21 mg RE g-1). Seventy four compounds were detected in B. microcarpum, 70 in B. brachyactis, 66 in B. sayai, and 51 in B. pinnatifolium. Quinic acid, chlorogenic acid, pantothenic acid, esculin, isoquercitrin, rutin, apigenin, and scopoletin were present in all the four species. This study showed that the four Bunium species are good sources of biologically active compounds with pharmaceutical and nutraceutical potential.


Assuntos
Apiaceae/química , Apiaceae/classificação , Amilases/antagonistas & inibidores , Amilases/metabolismo , Animais , Anti-Infecciosos/análise , Anti-Infecciosos/farmacologia , Antioxidantes/análise , Antioxidantes/farmacologia , Apigenina/análise , Apigenina/metabolismo , Ácido Clorogênico/análise , Ácido Clorogênico/farmacologia , Inibidores da Colinesterase/análise , Inibidores da Colinesterase/farmacologia , Enterobacter cloacae/efeitos dos fármacos , Enterobacter cloacae/metabolismo , Inibidores Enzimáticos/análise , Inibidores Enzimáticos/farmacologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Esculina/análise , Esculina/farmacologia , Glucosidases/antagonistas & inibidores , Glucosidases/metabolismo , Células HEK293 , Células Hep G2 , Humanos , Lipase/antagonistas & inibidores , Lipase/metabolismo , Camundongos , Testes de Sensibilidade Microbiana , Monofenol Mono-Oxigenase/antagonistas & inibidores , Monofenol Mono-Oxigenase/metabolismo , Ácido Pantotênico/análise , Ácido Pantotênico/farmacologia , Fenóis/análise , Fenóis/farmacologia , Extratos Vegetais/análise , Extratos Vegetais/farmacologia , Proteus mirabilis/efeitos dos fármacos , Proteus mirabilis/metabolismo , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/metabolismo , Quercetina/análogos & derivados , Quercetina/análise , Quercetina/farmacologia , Ácido Quínico/análise , Ácido Quínico/farmacologia , Células RAW 264.7 , Rutina/análise , Rutina/farmacologia
15.
J Exp Bot ; 70(20): 5559-5573, 2019 10 24.
Artigo em Inglês | MEDLINE | ID: mdl-31232453

RESUMO

Cassava (Manihot esculenta) is one of the most important staple food crops worldwide. Its starchy tuberous roots supply over 800 million people with carbohydrates. Yet, surprisingly little is known about the processes involved in filling of those vital storage organs. A better understanding of cassava carbohydrate allocation and starch storage is key to improving storage root yield. Here, we studied cassava morphology and phloem sap flow from source to sink using transgenic pAtSUC2::GFP plants, the phloem tracers esculin and 5(6)-carboxyfluorescein diacetate, as well as several staining techniques. We show that cassava performs apoplasmic phloem loading in source leaves and symplasmic unloading into phloem parenchyma cells of tuberous roots. We demonstrate that vascular rays play an important role in radial transport from the phloem to xylem parenchyma cells in tuberous roots. Furthermore, enzymatic and proteomic measurements of storage root tissues confirmed high abundance and activity of enzymes involved in the sucrose synthase-mediated pathway and indicated that starch is stored most efficiently in the outer xylem layers of tuberous roots. Our findings form the basis for biotechnological approaches aimed at improved phloem loading and enhanced carbohydrate allocation and storage in order to increase tuberous root yield of cassava.


Assuntos
Manihot/metabolismo , Floema/metabolismo , Raízes de Plantas/metabolismo , Transporte Biológico , Esculina/metabolismo , Regulação da Expressão Gênica de Plantas , Manihot/fisiologia , Floema/fisiologia , Proteínas de Plantas/metabolismo , Raízes de Plantas/fisiologia , Xilema/metabolismo , Xilema/fisiologia
16.
Methods Mol Biol ; 2014: 195-201, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31197797

RESUMO

Historically, the ability to measure the velocity of phloem sap in small seedlings and plants has been technically challenging. The phloem tissues are delicate, often flow is blocked entirely if perturbed. Furthermore, the depth that phloem sieve tubes are located within the plant has hindered many techniques. Previously published methods have lacked the spatial and temporal resolution required for measurements in small seedlings, are usually laborious or are not suited to in vivo studies. Here we describe a rapid, high-throughput method using the fluorescent coumarin glucoside esculin as a probe to measure the phloem transport velocity in the roots of young Arabidopsis seedlings.


Assuntos
Arabidopsis/metabolismo , Esculina/metabolismo , Floema/metabolismo , Plântula/metabolismo , Biomarcadores , Cumarínicos/metabolismo , Glucosídeos/metabolismo , Histocitoquímica
17.
Methods Mol Biol ; 2014: 253-266, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31197802

RESUMO

Sucrose transport across membranes requires the activity of transport proteins. Sucrose-specific SWEET proteins mediate sugar efflux out of the cytosol and SUC proteins catalyze the uptake of sucrose from the apoplast. Both transport processes are involved in phloem loading in source leaves as well as in the post-phloem pathway in sink tissues. An important step during the characterization of new sucrose transporters is to analyze their transport activity. This is usually achieved by heterologous expression of the respective gene in yeast cells or Xenopus oocytes and subsequent uptake measurements. However, in some cases, mistargeting to internal membranes or the lack of protein modifications and/or interaction partners in the heterologous system can interfere with uptake analyses. Therefore, a new in planta method was developed that is based on mesophyll protoplasts as expression system and the fluorescent sucrose analog esculin to monitor uptake activities by confocal microscopy. In this chapter we describe the design of constructs required to analyze sucrose transporters in protoplasts, the experimental setup of the protoplast-esculin assay, and the quantitative evaluation of the obtained data. The quantification of esculin uptake allows the application of the new assay to a variety of questions, e.g., by comparison of point mutants, splice variants, or transporters with and without interaction partners.


Assuntos
Bioensaio , Esculina/metabolismo , Protoplastos/metabolismo , Sacarose/metabolismo , Arabidopsis/metabolismo , Transporte Biológico , Microscopia Confocal , Floema/metabolismo , Folhas de Planta/metabolismo
18.
Eur J Pharmacol ; 857: 172453, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31202807

RESUMO

Aesculin, a natural product from the traditional and widely-used Chinese medicine named Cortex fraxini, has attracted attention as a novel therapeutic modulator of inflammation. However, little is known about its effect on ulcerative colitis (UC). This study aimed to investigate the protective effects and mechanisms of aesculin on colitis. The results showed that, few cytotoxicity of aesculin were shown in vivo and in the RAW264.7 macrophages, while aesculin significantly relieved the symptoms of DSS-induced colitis and restrained the expression of inflammatory factors including iNOS, IL-1ß, TNF-α in both peritoneal macrophages and colonic tissues from DSS-induced mice and RAW264.7 macrophages. Of note, aesculin attenuated the activity of NF-κB signaling while promoted the nuclear localization of PPAR-γ in both rectal tissues from DSS-induced mice and LPS-stimulated macrophages. These findings demonstrated that the protection of aesculin against ulcerative colitis might be due to its regulation on the PPAR-γ and NF-κB pathway. Thus, aesculin could serve as a potential therapeutic agent for the treatment of ulcerative colitis.


Assuntos
Colite/metabolismo , Colite/prevenção & controle , Sulfato de Dextrana/efeitos adversos , Esculina/farmacologia , NF-kappa B/metabolismo , PPAR gama/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Colite/induzido quimicamente , Colite/patologia , Citocinas/biossíntese , Citoproteção/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Camundongos , Células RAW 264.7
19.
Basic Clin Pharmacol Toxicol ; 125(3): 259-270, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30861618

RESUMO

The Keap1-Nrf2-ARE system serves as a premier defence mechanism to curb oxidative stress, which remains as one of the major causes of ageing and pathogenesis in various diseases. Nrf2 is the principal master regulator of the cellular defence system, and its activation remains the prospective therapeutic approach against chronic diseases. One of the recent strategies is to disrupt Keap1-Nrf2 protein-protein interaction (PPI) that alters the docking of Keap1 with Nrf2 by compounds occupying a position in the Keap1 blocking the interface with Nrf2. In this study, we made an attempt to identify the compounds with anticancer, antioxidant and anti-inflammatory properties to disrupt Keap1a/b-Nrf2 PPI through in silico molecular docking in zebrafish. The phylogenetic analysis of Keap1 proteins revealed the existence of orthologous Keap1-Nrf2-ARE system in lower vertebrates that includes zebrafish. The DGR domains of zebrafish Keap1a and Keap1b were modelled with Modeller 9.19 using Keap1 of Mus musculus (PDB ID:5CGJ) as template. Based on the docking calculations, top hit compounds were identified to disrupt both Keap1a and Keap1b interaction with Nrf2 which include quercetin 3,4'-diglucoside, flavin adenine dinucleotide disodium salt hydrate, salvianolic acid A, tunicamycin and esculin. The LC50 of esculin in 3 dpf zebrafish larvae is 5 mmol/L, and the qRT-PCR results showed that esculin significantly increased the transcription of Nrf2 target genes-Gstpi, Nqo1, Hmox1a and Prdx1 in 3 dpf zebrafish larvae. These potential hits could serve as safer Nrf2 activators due to their non-covalent disruption of Keap1-Nrf2 PPI and be developed into efficacious preventive/therapeutic agents for various diseases.


Assuntos
Antioxidantes/farmacologia , Descoberta de Drogas , Fator 2 Relacionado a NF-E2/agonistas , Domínios e Motivos de Interação entre Proteínas/efeitos dos fármacos , Proteínas de Peixe-Zebra/agonistas , Animais , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Embrião não Mamífero , Esculina/farmacologia , Dose Letal Mediana , Ligantes , Simulação de Acoplamento Molecular , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Filogenia , Ligação Proteica/efeitos dos fármacos , Domínios e Motivos de Interação entre Proteínas/genética , Transcrição Genética/efeitos dos fármacos , Peixe-Zebra , Proteínas de Peixe-Zebra/antagonistas & inibidores , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
20.
J Pharmacol Sci ; 139(3): 151-157, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30733181

RESUMO

Glucocorticoids are widely prescribed for lots of pathological conditions, however, can produce 'Cushingoid' side effects including central obesity, glucose intolerance, insulin resistance and so forth. Our study is intended to investigate the improving effects of coumarins on diabetogenic action of dexamethasone in vivo and in vitro and elucidate potential mechanisms. ICR mice treated with dexamethasone for 21 days exhibited decreased body weight, increased blood glucose and impaired glucose tolerance, which were prevented by fraxetin (40 mg/kg/day), esculin (40 mg/kg/day) and osthole (20 mg/kg/day), respectively. Esculin, fraxetin and osthole also could promote glucose uptake in normal C2C12 myotubes, and improve insulin resistance in myotubes induced by dexamethasone. Western blotting results indicated that esculin, fraxetin and osthole could boost Akt activation, stimulate GLUT4 translocation, thus alleviate insulin resistance. Esculin and osthole also could activate AMPK, thereby phosphorylate TBC1D1 at Ser237, and consequently ameliorate diabetogenic action of dexamethasone. Our study indicates coumarins as potential anti-diabetic candidates or leading compounds for drug development.


Assuntos
Glicemia/efeitos dos fármacos , Cumarínicos/farmacologia , Dexametasona/toxicidade , Músculo Esquelético/efeitos dos fármacos , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Western Blotting , Linhagem Celular , Cumarínicos/administração & dosagem , Dexametasona/administração & dosagem , Esculina/farmacologia , Glucocorticoides/administração & dosagem , Glucocorticoides/toxicidade , Hipoglicemiantes/administração & dosagem , Hipoglicemiantes/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Músculo Esquelético/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos
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